Prostate cancer (PCa) is the most commonly diagnosed malignancy and second leading cause of male cancer death in the U.S. While local disease can be effectively treated, there is no effective cure for non-organ confined disease. PCa is exquisitely dependent on androgen receptor (AR) signaling at all stages; therefore, ablation of AR activity (known as androgen deprivation therapy) is the first line of therapeutic intervention for disseminated tumors. While initially effective, recurrent, 'castrate-resistant' prostate cancers (CRPC) develop within 2-3 years as a result of inappropriately restored AR activity. Multiple pathways can induce recurrent AR activity but the most frequently observed alteration is enhanced accumulation of AR itself. Additionally, a small subset of cooperating oncogenes, especially c-Myc, has been shown to act in concert with AR to induce disease progression. Since no effective treatment has been identified for this incurable stage of disease, there is a significant need to determine the mechanisms that contribute to resurgent AR activity and CRPC, and to design effective new therapeutics for CRPC management. This application identifies the Ubiquitin-Specific Peptidase 22 (USP22) deubiquitylating enzyme as a combinatorial effector of both AR and c-Myc activity that is upregulated in PCa, and whose activity affords a novel, targetable molecule for therapeutic intervention. Several key preliminary findings demonstrate that: i. USP22 is significantly upregulated in prostate cancer; ii). USP22 regulates c-Myc transcriptional activation function; iii. USP22 is a potent regulator of AR accumulation and activity; iv. USP22 is required for accumulation of truncated receptors that drive CRPC formation; and v.) USP22 is critical for efficient AR-dependent PCa and CRPC proliferation. Based on these data, we hypothesize that USP22 is playing a role in tumor formation and transition to resistant disease. This hypothesis will be tested in two aims. First, the mechanism by which USP22 controls c-Myc and AR will be tested in cell models of both early stage and resistance disease. This approach involves depletion of USP22 expression and analysis of c-Myc and AR activity with respect to target gene transcription, protein stability/half- life, contribution of the proteasome, and cell cycle profile. Second, the relevance of USP22 towards PCa growth and progression will be determined. Orthotopic xenograft models will be employed to investigate the impact of USP22 depletion or overexpression in hormone sensitive and therapy resistant PCa. The outcome will be appraised based on tumor growth, intratumor proliferation, and serum PSA levels (representative of AR activity). This experimental design is a logical and efficient method to analyze the ability of USP22 to regulate the activity of two oncogenic signals that control PCa disease formation and progression. Together this proposal supports the hypothesis that USP22 is a master regulator of AR stability and c-Myc activity in PCa that could be effectively targeted in human disease for prevention and treatment of CRPC.